Cisplatinum Sensitivity of BRCA1-Mutated HCC1937 Breast Cancer Cells Is Linked to Impairment of Notch Signaling and Is Increased by γ-Secretase Inhibitors.

Abstract

Abstract BRCA1 plays a critical role in DNA-damage repair mechanisms elicited by cell exposure to anti-tumor agents. HCC1937 is a BRCA1-defective breast cancer cell line which discloses higher sensitivity to cisplatinum (CDDP) as compared to the BRCA1 full-length cDNA transfected clone, HCC1937/wtBRCA1. By cDNA microarray whole gene expression profile and Ingenuity Pathway Analysis (IPA) software, we found a differential modulation of Notch signaling after CDDP exposure in HCC1937 cell line as compared to BRCA1-reconstituted clone with a strong down-regulation of Notch1, 2 and 3 expression together with other genes involved in the Notch signaling network, including Hes1 and Jag1. Notch pathway was not affected by CDDP exposure in BRCA1-reconstituted cells. By Western Blot analysis, we next analyzed if the different trascriptional response of the two cell lines to CDDP occurred together with down-regulation of Notch protein in BRCA1-defective cells. We found enhanced expression of BRCA1 in BRCA1-transfected cells. We then analyzed the Notch 3 protein's expression in HCC1937 and HCC1937/wtBRCA1. By Western blot analysis, we demonstrated a significant reduction of Notch 3 protein's expression following CDDP exposure at the IC50 dose for 12, 18 and 24 hours, in HCC1937 as compared to HCC1937/wtBRCA1, consistently with gene expression results. In the aim of evaluating the role of Notch signaling in the regulation of the pharmacological response to CDDP in BRCA1-defective cells, we exposed HCC1937 cells to the pan-Notch inhibitor, γ-secretase inhibitor XII (GSI-XII), alone or in combination with CDDP, and we elvaluated, by MTT assay, in vitro growth effects on cell survival. We observed a time and dose dependent decrease of cell growth of HCC1937 following GSI-XII exposure as compared to BRCA1-reconstituted clone, with an IC50 between 25 and 35 microM as single agent, while HCC1937/wtBRCA1 where highly resistant to the drug. The combination of GSI-XII plus CDDP produced a significant sinergistic antitumor effect at 48 hours in HCC1937 which did not occurred in HCC1937/wtBRCA1. Our findings suggest that the high sensitivity of BRCA1-defective cells to CDDP exposure may be related not only to depression of the DNA-damage repair machinery but also to down-modulation of the Notch survival pathway. Moreover, our data indicate that the antitumor activity of CDDP is enhanced by combination with the Notch inhibitor GSI-XII suggesting that γ-secretase inhibitors may represent a potential and novel therapeutic tool in this specific setting. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 3128.